Having completed the primary structure of dogfish muscle lactate dehydrogenase and with the crystal structure known, it becomes possible to compare structural homologies which exist in other LDH's. By scanning a wide range of evolution, several unusual LDH's have been identified that differ significantly from the vertebrate ioenzymes. Consequently, this enzyme provides an excellent model for investigating the evolution of substrate specificity, subunit interaction, and also allosteric control at the level of the primary structure. To accomplish this, three particular enzymes have been selected for structural comparison. Lactate dehydrogenase from lobster tail is unique in its sigmoidal kinetics and also in the use of dimer-tetramer equilibrium as a control mechanism. Horseshoe crab has an LDH that is specific for D-lactate and is also dimeric. Finally, tetrameric LDH from Lactobacillus casei is allosterically modulated by fructose-1,6-diphosphate. Therefore, these three enzymes, while all maintaining the same subunit molecular weight, have developed some unique properties which distinguish them markedly from the vertebrate LDH isoenzymes and provide an excellent system for studying the diverse evolution of specific regions of the LDH molecule. A method has been developed for isolating rapidly some of the peptides that compose the active site regions of LDH including the "loop" region that shows major conformational changes following coenzyme binding and part of the substrate binding site. Characterization of these peptides so far has indicated clear homologies in these active site regions of LDH from lobster tail and L. casei whereas the sequence homologies with horseshoe crab are much less obvious. After characterizing these active site peptides, the complete amino acid sequence of each enzyme will be elucidated. Sequencing will be carried out by the manual dansyl-Edman technique as well as by automated solid-phase sequencing procedures.